Project Summary: Mechanistically Linking Insulin Action and the Thermic Effect of Food Positive energy balance leading to ectopic fat deposition has been shown to play a major role in the pathogenesis of insulin resistance and type 2 diabetes, a disease that costs the United States healthcare system over 300 billion dollars a year. One important component of energy expenditure is the thermic effect of food (TEF), which represents the increase in heat production and oxygen consumption following nutrient intake. However, the mechanistic link between nutrient intake and energy expenditure remains poorly understood. Insulin stimulated cleavage of the TUG protein has been shown to regulate glucose uptake and vasopressin inactivation in vitro and in vivo. Unexpectedly, mice with constitutive and unregulated cleavage of TUG proteins exhibit significantly increased energy expenditure that is associated with transcriptional induction of proteins that mediate futile cycles of metabolic substrates. Conversely, mice with muscle specific deletion of TUG have decreased expression of thermogenic proteins. This proposal will test the hypothesis that the TUG C-terminal cleavage product regulates energy expenditure by localizing to the nucleus and modulating gene expression. The first aim is to study the mechanism of increased energy expenditure in transgenic mice with constitutive TUG cleavage. The dynamics of TUG C-terminal product localization and how it is affected in insulin resistant tissues will be studied in complementary mouse strains. Interaction partners of the TUG C-terminus within the nucleus will be identified using unbiased screens and verified in both mice and cell lines. The second aim is to understand how degradation of the TUG C-terminus is regulated by the N-end rule, which is well described to regulate the half-life of cleavage products. Tagged fusion proteins, siRNA, and in vivo characterization will be used to determine how the abundance of TUG may be regulated. Overall, this proposal will elucidate a novel link between insulin signaling and energy expenditure and provide the groundwork for further studies on how this process may be regulated. Such studies will also allow for the identification of potential sites of pharmacologic intervention. Finally, the proposal includes a comprehensive training plan that will provide important new learning experiences to support the applicant?s development as a physician-scientist who is able to integrate scientific work and knowledge of diabetes with novel clinically relevant research.